Quantitation of protein interactions in cancer cells

描述(申请人提供)：蛋白质相互作用是生物系统中蛋白质功能的关键决定因素。尽管定量蛋白质相互作用信息可能对癌症研究的所有领域具有潜在的作用，但对原生生命系统内的蛋白质相互作用进行无偏见或大规模量化是当今技术无法满足的挑战。目前尚不存在在天然细胞、患者样本或组织内大规模识别和量化蛋白质相互作用的能力。提高量化蛋白质相互作用的能力将对了解癌症、转移和抗癌耐药性的发展产生重大影响。该项目旨在开发和应用先进的蛋白质相互作用报告(PIR)技术与癌细胞进行定量交联。细胞培养中氨基酸的稳定同位素标记(SILAC)将与PIR技术相结合，以实现对细胞中蛋白质水平和蛋白质相互作用的定量。这些能力将应用于顺铂、紫杉醇和SN-38耐药癌细胞，以允许量化与药物敏感癌细胞的相互作用。该项目将提供第一个关于癌细胞中蛋白质相互作用的相对定量数据，以及第一个与耐药相关的蛋白质相互作用水平上的功能调节的无偏见测量。 公共卫生相关性：癌症治疗中的耐药性是治疗失败的主要原因，在更好地了解支持耐药性的功能调控之前，它可能仍然是导致癌症患者死亡的主要因素。所有细胞的功能调节都是通过改变蛋白质的丰度、定位、相互作用和拓扑特征来实现的。该项目将开发和应用先进技术，以帮助可视化已获得耐药性的癌细胞功能调节的变化。

DESCRIPTION (provided by applicant): Protein interactions are key determinants of protein function in biological systems. Despite the potential that quantitative protein interaction information could have for all areas of cancer research, unbiased or large-scale quantitation of protein interactions within native living systems is a challenge that is unmet by today's technology. The capacity to identify and quantitate protein interactions on a large-scale within native cells, patient samples, or tissues does not currently exist. Improved capabilities to quantitate protein interactions will have a major impact on the understanding of cancer, metastasis and the development of anti-cancer drug resistance. This project aims to develop and apply quantitative cross-linking with cancer cells with advanced Protein Interaction Reporter (PIR) technology. Stable Isotope Label of Amino acids in Cell culture (SILAC) will be combined with PIR technology to allow quantitation of protein levels and protein interactions in cells. These capabilities will be applied to cisplatin-, taxol-, and SN-38 resistant cancer cells to allow quantitation of interactions relative to drug sensitive cancer cells. This project will provide the first relative quantitation data on protein interactions in cancer cells and the first unbiased measurements of functional regulation at the protein interaction level relevant to drug resistance. PUBLIC HEALTH RELEVANCE: Drug resistance in cancer treatment is the primary reason for therapy failure and will likely remain a primary factor leading to cancer patient death until functional regulation that supports drug resistance can be better understood. Functional regulation in all cells is achieved by changes in protein abundance, localization, interactions and topological features. This project will develop and apply advanced technology to help visualize changes in functional regulation in cancer cells that have acquired drug resistance.